The in vivo mutation assay using the X-linked phosphatidylinositol glycan class A gene (Pig-a in rodents, PIG-A in humans) is a promising tool for evaluating the mutagenicity of chemicals. Approaches for measuring Pig-a mutant cells have focused on peripheral red blood cells (RBCs) and reticulocytes (RETs) from rodents. The recently developed PIGRET assay is capable of screening >1x106 RETs for Pig-a mutants by concentrating RETs in whole blood prior to flow cytometric analysis. Additionally, due to the characteristics of erythropoiesis, the PIGRET assay can potentially detect increases in Pig-a mutant frequency (MF) sooner after exposure compared with a Pig-a assay targeting total RBCs (RBC Pig-a assay). In order to test the merits and limitations of the PIGRET assay as a short-term genotoxicity test, an interlaboratory trial involving 16 laboratories was organized by the Mammalian Mutagenicity Study Group of the Japanese Environmental Mutagenicity Society (MMS/JEMS). First, the technical proficiency of the laboratories and transferability of the assay were confirmed by performing both the PIGRET and RBC Pig-a assays on rats treated with single doses of N-nitroso-N-ethylurea. Next, the collaborating laboratories used the PIGRET and RBC Pig-a assays to assess the mutagenicity of a total of 24 chemicals in rats, using a single treatment design and mutant analysis at 1, 2, and 4 weeks after the treatment. Thirteen chemicals produced positive responses in the PIGRET assay; three of these chemicals were not detected in the RBC Pig-a assay. Twelve chemicals induced an increase in RET Pig-a MF beginning 1 week after dosing, while only 3 chemicals positive for RBC Pig-a MF produced positive responses 1 week after dosing. Based on these results, we conclude that the PIGRET assay is useful as a short-term test for in vivo mutation using a single-dose protocol.

中文翻译：

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PIGRET 试验是一种测量网织红细胞中 Pig-a 基因突变的方法，作为短期体内基因毒性试验是可靠的：MMS/JEMS 合作研究的摘要，涉及 16 个实验室使用 24 种化学品。

使用 X 连锁磷脂酰肌醇聚糖 A 类基因（啮齿动物中的 Pig-a，人类中的 PIG-A）进行的体内突变测定是评估化学品致突变性的一种很有前景的工具。测量 Pig-a 突变细胞的方法主要集中在啮齿动物的外周血红细胞 (RBC) 和网织红细胞 (RET)。最近开发的 PIGRET 检测能够通过在流式细胞术分析之前在全血中浓缩 RET 来筛选 >1x106 RET 的 Pig-a 突变体。此外，由于红细胞生成的特性，与针对总红细胞的 Pig-a 检测（RBC Pig-a 检测）相比，PIGRET 检测可以在暴露后更快地检测到 Pig-a 突变频率 (MF) 的增加。为了测试 PIGRET 试验作为短期遗传毒性试验的优点和局限性，日本环境致突变性学会 (MMS/JEMS) 的哺乳动物致突变性研究组组织了一项涉及 16 个实验室的实验室间试验。首先，通过对接受单剂量 N-亚硝基-N-乙基脲治疗的大鼠进行 PIGRET 和 RBC Pig-a 检测，证实了实验室的技术熟练程度和检测的可转移性。接下来，合作实验室使用 PIGRET 和 RBC Pig-a 检测，使用单一治疗设计和治疗后 1、2 和 4 周的突变分析来评估总共 24 种化学物质对大鼠的致突变性。13 种化学物质在 PIGRET 试验中产生了阳性反应；在 RBC Pig-a 检测中未检测到其中三种化学物质。在给药后 1 周开始，12 种化学物质诱导了 RET Pig-a MF 的增加，而只有 3 种对 RBC Pig-a MF 呈阳性的化学物质在给药 1 周后产生了阳性反应。基于这些结果，我们得出结论，PIGRET 检测可用作使用单剂量方案进行体内突变的短期测试。